1. Evaluation of the Efficacy of Infectious
Bursal Disease Virus Multivalent Virus-
Like-Particle Antigens in Enzyme-Linked
Immunosorbent Assays
Kimberly Carter
Mentor: Dr. Daral Jackwood
Food Animal Health Research Program

2. Infectious Bursal Disease Virus
• Highly contagious
• Targets immature B
lymphocytes
• Damages bursa of Fabricius
and other lymphoid organs
• Immunosuppression

3. Infectious Bursal
Disease Virus
Serotype 1
Classic Variant
Serotype 2
There are many different variant strains!

4. Antigenic Drift
Variations
in P loops
New variant
strains
VP2
Protein

5. Significance of Antigenic Drift
• Antibodies against classic strains cannot
neutralize variant strains and vice versa
• Some antibodies against variant strains
cannot be detected by an ELISA

6. Potential of multivalent virus-like
particles
• Co-expression of pVP2
and VP3 produces
virus-like particles
(VLPs)
• Co-expression of variant
and classic pVP2s
produces multivalent
VLPs

7. Objective: Compare the efficacy of multivalent VLP antigens in
ELISAs to the efficacy of antigens found in commercially-
produced ELISAs in detecting classic and variant IBDV antibodies
Hypothesis: Multivalent VLP antigens in ELISAs will yield more
positive results using known chicken anti-IBDV sera than
commercial ELISA kits

8. Materials and Methods: Multivalent VLP
Antigen Production
VP3 pVP2 Classic pVP2 Variant
Sf9 insect cells
IBDV Multivalent VLP antigens

9. Materials and Methods: ELISA
96 well flat-bottomed plate
IBDV multivalent
VLP antigens
96 well flat-bottomed plate
Various chicken
anti-IBDV serum
samples
96 well flat-bottomed plate
Peroxidase-labeled
goat anti-chicken
immunoglobulin G
(conjugate)

10. ELISA (continued)
96 well flat-bottomed plate
ABTS substrate
added and
converted to a
detectable product
by the conjugate
96 well flat-bottomed plate
Stop solution added
after 15 minutes to
stop color
development

11. Materials and Methods: Optical Density
Measurement
• Plates were read on an ELISA reader at 405 nm

12. Results:
FD181 VP3, pf33 pVP2, T1 pVP2, and pp34 pVP2 antigen
0.000
0.100
0.200
0.300
0.400
0.500
0.600
0.700
0.800
0.900
1.000
OpticalDensity(OD405)
100
200
400
800
1600
3200
6400
Control
Vertical bars represent the mean of 12 serum samples and their dilutions from 1:100-1:6400.
Horizontal bar represents 2 standard deviations above the negative control sera.

13. Results:
FD181 VP3, Mo195 pVP2, T1 pVP2, and pp34 pVP2
0.000
0.100
0.200
0.300
0.400
0.500
0.600
0.700
0.800
0.900
1.000
1.100
OpticalDensity(OD405)
100
200
400
800
1600
3200
6400
Control
Vertical bars represent the mean of 12 serum samples and their dilutions from 1:100-1:6400.
Horizontal bar represents 2 standard deviations above the negative control sera.

14. Results:
FD181 VP3, Mo195 pVP2, pf33 pVP2, T1 pVP2, and pp34 pVP2
0.000
0.100
0.200
0.300
0.400
0.500
0.600
0.700
0.800
0.900
1.000
OpticalDensity(OD405)
100
200
400
800
1600
3200
6400
Control
Vertical bars represent the mean of 12 serum samples and their dilutions from 1:100-1:6400.
Horizontal bar represents 2 standard deviations above the negative control sera.

15. Results:
FD181 VP3, Mo195 pVP2, and pp34 pVP2
0.000
0.100
0.200
0.300
0.400
0.500
0.600
0.700
0.800
OpticalDensity(OD405)
100
200
400
800
1600
3200
6400
Control
Vertical bars represent the mean of 12 serum samples and their dilutions from 1:100-1:6400.
Horizontal bar represents 2 standard deviations above the negative control sera.

16. Conclusions and Future Directions
• Effective in yielding positive results at a dilution of
1:100
– Not nearly as effective at higher dilutions
• FD181 VP3, Mo195 pVP2, and pp34 pVP2 antigen
appeared to be most effective
– Unexpected
• Commercially-produced ELISAs need to be conducted
– Are ELISAs containing multivalent VLP antigens
more effective than commercial ELISA kits?

17. Multivalent vs. Monovalent VLPs
• IBDV monovalent VLP antigens yielded more
positive results at higher dilutions
– Positive results at a 1:200 dilution in a majority of serum
samples
– Multiple positive results at a dilution of 1:400
• Suggests monovalent VLP antigens are more
effective than multivalent VLP antigens

18. References
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19. Acknowledgements
• Dr. Daral Jackwood
• Fellow colleagues in Dr.
Jackwood’s lab
• All personnel in the Food
Animal Health Research
Program
• All who make ORIP
possible

20. Questions?